title: The human topoisomerase 1B Arg634Ala mutation results in camptothecin resistance and loss of inter-domain motion correlation.
abstract: Human topoisomerase 1B, the unique target of the natural anticancer compound camptothecin, catalyzes the unwinding of supercoiled DNA by introducing transient single strand nicks and providing covalent protein-DNA adducts. The functional properties and the drug reactivity of the single Arg634Ala mutant have been investigated in comparison to the wild type enzyme. The mutant is characterized by an identical relaxation and cleavage rate but it displays resistance to camptothecin as indicated by a viability assay of the yeast cells transformed with the mutated protein. The mutant also displays a very fast religation rate that is only partially reduced by the presence of the drug, suggesting that this is the main reason for its resistance. A comparative analysis of the structural-dynamical properties of the native and mutant proteins by molecular dynamics simulation indicates that mutation of Arg634 brings to a loss of motion correlation between the different domains and in particular between the linker and the C-terminal domain, containing the catalytic tyrosine residue. These results indicate that the loss of motion correlation and the drug resistance are two strongly correlated events.
authors.affiliation: Department of Biology and Interuniversity Consortium, National Institute Biostructure and Biosystem (INBB), University of Rome Tor Vergata, Via Della Ricerca Scientifica, Rome 00133, Italy.
authors.forename: Ilda
authors.initials: I
authors.lastname: D'Annessa
authors.forename: Cinzia
authors.initials: C
authors.lastname: Tesauro
authors.forename: Zhenxing
authors.initials: Z
authors.lastname: Wang
authors.forename: Barbara
authors.initials: B
authors.lastname: Arnò
authors.forename: Laura
authors.initials: L
authors.lastname: Zuccaro
authors.forename: Paola
authors.initials: P
authors.lastname: Fiorani
authors.forename: Alessandro
authors.initials: A
authors.lastname: Desideri
mesh_terms: D019943:Amino Acid Substitution; D002166:Camptothecin / Q000737:chemistry* / Q000494:pharmacology; D004264:DNA Topoisomerases, Type I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D006801:Humans; D056004:Molecular Dynamics Simulation; D020125:Mutation, Missense; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D059004:Topoisomerase I Inhibitors / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D059004:Topoisomerase I Inhibitors; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human; D002166:Camptothecin
keywords: BSA; CPT; Camptothecin; DMSO; DTT; EDTA; EGTA; Linker domain; Molecular dynamics; PMSF; Single mutation; TBE; TPT; Topoisomerase; bovine serine albumin; camptothecin; dimethyl sulfoxide; dithiothreitol; ethylene glycol tetraacetic acid; ethylenediaminetetraacetic acid; hTop1B; human topoisomerase 1B; phenylmethanesulfonylfluoride; topotecan; tris-borate-EDTA